Stroke remains the third most common cause of death in the industrial world, ranking behind ischemic heart disease and cancer. Strokes are responsible for about 300,000 deaths annually in the United States and about 11,000 deaths annually in Austria. Strokes are also a leading cause of hospital admissions and long-term disabilities. Accordingly, the socioeconomic impact of stroke and its attendant burden on society is practically immeasurable.
"Stroke" is defined by the World Health Organization as a rapidly developing clinical sign of focal or global disturbance of cerebral function with symptoms lasting at least 24 hours. Strokes are also implicated in deaths where there is no apparent cause other than an effect of vascular origin.
Strokes are typically caused by blockages or occlusions of the blood vessels to the brain or within the brain. With complete occlusion, arrest of cerebral circulation causes cessation of neuronal electrical activity within seconds. Within a few minutes after the deterioration of the energy state and ion homeostasis, depletion of high energy phosphates, membrane ion pump failure, efflux of cellular potassium, influx of sodium chloride and water, and membrane depolarization occur. If the occlusion persists for more than five to ten minutes, irreversible damage results. With incomplete ischemia, however, the outcome is difficult to evaluate and depends largely on residual perfusion and the availability of oxygen. After a thrombotic occlusion of a cerebral vessel, ischemia is rarely total. Some residual perfusion usually persists in the ischemic area, depending on collateral blood flow and local perfusion pressure.
Cerebral blood flow can compensate for drops in mean arterial blood pressure from 90 to 60 mm Hg by autoregulation. This phenomenon involves dilatation of downstream resistant vessels. Below the lower level of autoregulation (about 60 mm Hg), vasodilatation is inadequate and the cerebral blood flow falls. The brain, however, has perfusion reserves that can compensate for the fall in cerebral blood fall. This reserve exists because under normal conditions only about 35% of the oxygen delivered by the blood is extracted. Therefore, increased oxygen extraction can take place, provided that normoxia and normocapnea exist. When distal blood pressure falls below approximately 30 mm Hg, the two compensatory mechanisms (autoregulation and perfusion reserve) are inadequate to prevent failure of oxygen delivery.
As flow drops below the ischemic threshold of 23 ml/100 g/minute, symptoms of tissue hypoxia develop. Severe ischemia may be lethal. When the ischemia is moderate, it will result in "penumbra." In the neurological context, penumbra refers to a zone of brain tissue with moderate ischemia and paralyzed neuronal function, which is reversible with restoration of adequate perfusion. The penumbra forms a zone of collaterally perfused tissue surrounding a core of severe ischemia in which an infarct has developed. In other words, the penumbra is the tissue area that can be saved, and is essentially in a state between life and death.
When a clot is degraded and the blood flow to the penumbra is restored, a phenomenon known as reperfusion injury can occur. Portions of the injured tissue in the penumbra can be killed or further injured by the re-entry of oxygen or other substances into the area affected by the ischemia. In view of this phenomenon, the extent of tissue damage resulting from ischemia is determined both by the time required to achieve opening of an occluded vessel and by the series of reactions that follow as a result of reperfusion and the re-entry of oxygen to the affected tissue.
Although an ischemic event can occur anywhere in the vascular system, the carotid artery bifurcation and the origin of the internal carotid artery are the most frequent sites for thrombotic occlusions of cerebral blood vessels, which result in cerebral ischemia. The symptoms of reduced blood flow due to stenosis or thrombosis are similar to those caused by middle cerebral artery disease. Flow through the ophthalmic artery is often affected sufficiently to produce amaurosis fugax or transient monocular blindness. Severe bilateral internal carotid artery stenosis may result in cerebral hemispheric hypoperfusion. This manifests with acute headache ipsilateral to the acutely ischemic hemisphere. Occlusions or decrease of the blood flow with resulting ischemia of one anterior cerebral artery distal to the anterior communicating artery produces motor and cortical sensory symptoms in the contralateral leg and, less often, proximal arm. Other manifestations of occlusions or underperfusion of the anterior cerebral artery include gait ataxia and sometimes urinary incontinence due to damage to the parasagittal frontal lobe. Language disturbances manifested as decrease spontaneous speech may accompany generalized depression of psychomotor activity.
Most ischemic strokes involve portions or all of the territory of the middle cerebral artery with emboli from the heart or extracranial carotid arteries accounting for most cases. Emboli may occlude the main stem of the middle cerebral artery, but more frequently produce distal occlusion of either the superior or the inferior branch. Occlusions of the superior branch cause weakness and sensory loss that are greatest in the face and arm. Occlusions of the posterior cerebral artery distal to its penetrating branches cause complete contralateral loss of vision. Difficulty in reading (dyslexia) and in performing calculations (dyscalculia) may follow ischemia of the dominant posterior cerebral artery. Proximal occlusion of the posterior cerebral artery causes ischemia of the branches penetrating to calamic and limbic structures. The clinical results are hemisensory disturbances that may chronically change to intractable pain of the defective site (thalamic pain).
A significant event in cerebral ischemia is known as the transient ischemic attack ("TIA"). A TIA is defined as a neurologic deficit with a duration of less than 24 hours. The TIA is an important sign of a ischemic development that may lead to cerebral infarction. Presently, no ideal treatment for TIA exists, and there are no generally accepted guidelines as to whether medical or surgical procedures should be carried out in order to reduce the incidence of stroke in subjects with TIA.
The etiology of TIA involves hemodynamic events and thromboembolic mechanisms. Because most TIAs resolve within one hour, a deficit that lasts longer is often classified as presumptive stroke and is, accordingly, associated with permanent brain injury. Therefore, computed tomographic brain scans are used to search for cerebral infarction in areas affected by TIAs lasting longer than several hours. In sum, the relevant clinical distinction between a TIA and a stroke is whether the ischemia has caused brain damage, which is typically classified as infarction or ischemic necrosis. Subjects with deteriorating clinical signs might have stroke in evolution or are classified as having progressive stroke. In this clinical setting, clot propagation is possibly an important factor in disease progression.
There are a myriad of other diseases caused by or associated with ischemia. Vertebrobasilar ischemia is the result of the occlusion of the vertebral artery. Occlusion of the vertebral artery and interference with flow through the ipsilateral posterior inferior cerebellar artery causes lateral medullary syndrome, which has a symptomology including vertigo, nausea, vomiting nystagmus, ipsilateral ataxia and ipsilateral Herner's syndrome. Vertebrobasilar ischemia often produces multifocal lesions scattered on both sides of the brain stem along a considerable length. Except for cerebellar infarction and the lateral medullary syndrome, the clinical syndromes of discrete lesions are thus seldom seen in pure form. Vertebrobasilar ischemia manifests with various combinations of symptoms such as dizziness, usually vertigo, diplopia, facial weakness, ataxia and long tract signs.
A basilar artery occlusion produces massive deficits. One of these deficits is known as the "locked in state." In this condition, paralysis of the limbs and most of the bulbar muscle leaves the subject only able to communicate by moving the eyes or eyelids in a type of code. Occlusion of the basilar apex or top of the basilar is usually caused by emboli that lodge at the junction between the basilar artery and the two posterior cerebral arteries. The condition produces an initial reduction in arousal followed by blindness and amnesia due to an interruption of flow into the posterior cerebral arteries as well as abnormalities of vertical gaze and pupillary reactivity due to tegmental damage.
Venous occlusion can cause massive damage and death. This disease is less common than arterial cerebral vascular disease. As with ischemic stroke from arterial disease, the primary mechanism of brain damage is the reduction in capillary blood flow, in this instance because of increased outflow resistance from the blocked veins. Back transmission of high pressure into the capillary bed usually results in early brain swelling from edema and hemorrhagic infarction in subcortical white matter. The most dangerous form of venous disease arises when the superior sagittal sinus is occluded. Venous occlusion occurs in association with coagulation disorders, often in the purpural period, or in subjects with disseminated cancers or contagious diseases. If anticoagulant therapy is not initiated, superior sagittal sinus occlusion has a mortality rate of 25-40%.
Brief diffuse cerebral ischemia can cause syncope without any permanent sequelae. Prolonged diffuse ischemia in other organs has devastating consequences. The most common cause is a cardiac asystole or other cardiopulmonary failures, including infarction. Aortic dissection and global hypoxia or carbon monoxide poisoning can cause similar pictures. Diffuse hypoxia/ischemia typically kills neurons in the hippocampus, cerebellar Purkinje cells, striatum or cortical layers. Clinically, such a diffuse hypoxia/ischemia results in unconsciousness and in coma, followed in many instances by a chronic vegetative state. If the subject does not regain consciousness within a few days, the prognosis for the return of independent brain functions becomes very poor.
Hyperviscosity syndrome is another disease related to blood flow and ischemia. Cerebral blood flow is inversely related to blood viscosity. The latter is directly proportional to the number of circulating red and white cells, the aggregation state, the number of platelets and the plasma protein concentration. Blood flow is inversely proportional to the deformability of erythrocytes and blood velocity (shear rate). Subjects with hyperviscosity syndrome can present either with focal neurologic dysfunction or with diffuse or multifocus signs or symptoms including headache, visual disturbances, cognitive impairments or seizures.
There are a number of substances involved in clot formation and lysis. Plasminogen, also known as gluplasminogen, and plasmin are two of the primary substances involved in lysis.
Plasminogen is a protein, composed of 791 amino acids, that circulates in human plasma at a concentration of about 200 .mu.g/ml. Plasminogen is the zymogen form of the fibrinolytic enzyme, plasmin, which has broad substrate specificity and is ultimately responsible for degrading blood clots. For the most part, fibrin proteolysis is mediated by the generation of plasmin within a fibrin clot from the plasminogen trapped within the clot. Fredenburgh & Nesheim, J. Biol. Chem. 267: 26150-56 (1992).
Plasminogen has five kringle domains within its amino-terminal heavy chain region that exhibit lysine-binding sites for recognition of lysine residues in fibrin. Plasminogen-plasmin conversion, both within a clot and at its surface, is facilitated by the affinity of tissue plasminogen activator ("t-PA") for fibrin, which results in a fibrin-dependent t-PA-induced plasminogen activation. Fredenburgh, loc. cit.
Once initiated, fibrinolysis results in several positive feedback reactions. For instance, plasmin-catalyzed cleavage of fibrin generates carboxy-terminal lysine residues, which in turn provide additional binding sites for both t-PA and plasminogen. This also facilitates plasmin-catalyzed conversion of glu-plasminogen to lys-plasminogen by a specific cleavage, which is a pre-activation step. Shih & Hajjar, P.S.E.B.M. 202: 258-64 (1993). In the literature, the term "lys-plasminogen" refers to forms of plasminogen where the N-terminal amino acid is lysine, valine or methionine. Nieuwenhuizen et al., Eur. J. Biochem. 174: 163-69 (1988). The conversion activity reflects a positive feedback reaction, because lys-plasminogen is a considerably better substrate than glu-plasminogen for both t-PA and urokinase, which may be caused by the enhanced affinity of lys-plasminogen for fibrin. The ratio of the Kcat/Km for t-PA-catalyzed activation of lys-plasminogen exceeds that of glu-plasminogen by about a factor of ten. Fredenburgh, loc. cit.
Fibrinolysis has been previously shown to be accelerated by the addition of lys-plasminogen in vivo and in vitro. In addition, when the use of exogenous lys-plasminogen was compared to the use of exogenous glu-plasminogen in similar experiments, 0.08 .mu.mol lys-plasminogen produced the same degree of enhanced fibrinolysis as 0.67 .mu.mol glu-plasminogen. Therefore, while both forms of plasminogen shorten the time for fibrinolysis, lys-plasminogen is about eight times more potent in this regard than glu-plasminogen. See Fredenburgh, supra.
Previous studies of lys-plasminogen have not implicated this protein for use in treatment of reperfusion injury. The conventional treatment for reperfusion injury is Flunarizine, which is only effective when administered prophylactically.